title: Design and control of acetylcholine receptor conformational change.
abstract: Allosteric proteins use energy derived from ligand binding to promote a global change in conformation. The "gating" equilibrium constant of acetylcholine receptor-channels (AChRs) is influenced by ligands, mutations, and membrane voltage. We engineered AChRs to have specific values of this constant by combining these perturbations, and then calculated the corresponding values for a reference condition. AChRs were designed to have specific rate and equilibrium constants simply by adding multiple, energetically independent mutations with known effects on gating. Mutations and depolarization (to remove channel block) changed the diliganded gating equilibrium constant only by changing the unliganded gating equilibrium constant (E(0)) and did not alter the energy from ligand binding. All of the tested perturbations were approximately energetically independent. We conclude that naturally occurring mutations mainly adjust E(0) and cause human disease because they generate AChRs that have physiologically inappropriate values of this constant. The results suggest that the energy associated with a structural change of a side chain in the gating isomerization is dissipated locally and is mainly independent of rigid body or normal mode motions of the protein. Gating rate and equilibrium constants are estimated for seven different AChR agonists using a stepwise engineering approach.
authors.affiliation: Department of Physiology and Biophysics, 309 Cary Hall, State University of New York, Buffalo, NY 14214, USA.
authors.forename: Snehal V
authors.initials: SV
authors.lastname: Jadey
authors.forename: Prasad
authors.initials: P
authors.lastname: Purohit
authors.forename: Iva
authors.initials: I
authors.lastname: Bruhova
authors.forename: Timothy M
authors.initials: TM
authors.lastname: Gregg
authors.forename: Anthony
authors.initials: A
authors.lastname: Auerbach
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002138:Calibration; D018679:Cholinergic Agonists / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects; D007700:Kinetics; D051379:Mice; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D017433:Protein Structure, Secondary; D011759:Pyrrolidines / Q000494:pharmacology; D011950:Receptors, Cholinergic / Q000737:chemistry* / Q000378:metabolism
chemical_list: D018679:Cholinergic Agonists; D050505:Mutant Proteins; D011759:Pyrrolidines; D011950:Receptors, Cholinergic; D000109:Acetylcholine